Location: Subtropical Horticulture Research2012 Annual Report
1a. Objectives (from AD-416):
1. To screen ARS germplasm collection maintained as the SHRS in Miami for tolerance/resistance to the disease. 2. To develop 10 to 30 thousand usable Simple Sequence Repeat (SSR) and Single Nucleotide Polymorphism (SNP) markers to saturate our current genetic linkage maps and to use for association mapping to locate genes involved with tolerance/resistance.
1b. Approach (from AD-416):
Resistance/tolerance to Laurel Wilt will be evaluated by screening maternal half-sib families from each of the ~300 germplasm accessions in the collection. The seeds will be harvested over the next two seasons (June-November 2010 and 2011) and germinated at the USDA-ARS station in Ft. Pierce, FL. Our objectives are to screen 100 seedlings from each accession and to estimate the heritability of the disease resistance/tolerance from maternal parents conferring tolerance. The plants will be planted using ~four ha of the USHRL farm or alternatively the adjacent IFAS farm. The plants will be inoculated with Raffaelea lauricola in the field and scored for tolerance to the pathogen. Tolerant genotypes will be propagated for further testing. A subset of the 30,000 seedlings will be evaluated using microsatellite and SNP markers developed from NGS. Using the SNP chip array will allow us to map 15,000 loci in each individual in a single evaluation. Using this information, we will perform association mapping to determine areas of the avocado genome involved with tolerance to laurel wilt. Molecular analysis will be used to find associations between molecular markers and the tolerance as a tool for enhanced screening. Using next generation sequencing (NGS), thousands of SSR and SNP markers can be easily identified and used for increased map saturation and association genetics thus facilitating QTL identification. We will generate 10 x 454 coverage of the cultivar ‘Hass’ and 50x Illumina coverage for the other three parents of our mapping populations, ‘Bacon’, ‘Simmonds’, and ‘Tonnage’. The identification of 300,000 new SNP markers has been accomplished using this procedure with cacao (Theobroma cacao L.). The SNP markers will be used to develop an Illumina 30K SNP iSelect array.
3. Progress Report:
This project is related to the inhouse objective: To investigate the genetic basis of important horticultural traits such as disease resistance and to select improved germplasm. Molecular markers have been developed for genetic diversity analysis and for the production of molecular genetic linkage maps. Families of avocado, mango, and jackfruit have been produced that should allow the mapping of Quantitative Trait Loci (QTL) involved with disease resistance, fruit quality, and yield. A medium to high-throughput single nucleotide polymorphism (SNP) assay platform was obtained through supplementary funding to genotype the open-pollinated progeny from germplasm accessions that have been planted at the USHRL, Fort Pierce, FL. Laurel wilt has been observed in avocado production orchards in Miami-Dade County within 20 miles of the SHRS station. Open pollinated seed of Mexican accessions has been obtained from University of California, Riverside collaborators for germination and planting at both SHRS and USHRL to identify laurel wilt resistant individuals and to estimate heritability of laurel wilt resistance.